Construction Of Structurally Novel Steroidal Heterocycles And Their Antitumor Activity Evaluation

Steroids, an important class of polycyclic molecules, play vital roles in maintaining normal physiological functions and in disease treatment. Steroid-based new drug discovery has long been pursued. In last decades, significant progress has been witnessed and over 300 steroidal compounds are currently being used in clinic for the treatment of diseases. The construction of structurally novel and diverse steroids and further biological assessment are of high interest in the field of steroid-based new drug discovery. Steroid-based chemical modifications focus on the construction of steroidal heterocycles. The incorporation of heterocycles into the steroid nucleus is capable of increasing the diversity, novelty and complexity of steroids, as well as always endows the steroidal heterocycles with new and/or improved biological activites. The construction of steroidal heterocycles will provide basis for steroid-based new drug discovery. The work described herein includes the followings:1. Based on the base-promoted cascade Aldol/cycloaddition sequence between the steroidal dicyano alkenes and isatins, we synthesized two series of structurally novel steroidal spiro-pyranyl oxindoles. One C-C single bond, one C-O bond, one quaternary stereogenic center, and one spirocyclic scaffold were formed in this one-pot reaction. Of these synthesized compounds, compound Ⅱ-6a showed broad and good antiproliferative activity toward different cancer cell lines and low toxicity toward normal cells. Further mechanistical studies indicated that compound Ⅱ-6a induced cancer cell death through ROS-mediated mechanisms. In vivo studies suggested that compound Ⅱ-6a had no remarkable toxicity toward the tested animals even at high doses and exerted favorable in vivo antitumor activity.2. Based on the steroidal unsaturated ketones, the [3+2] cycloaddition was then employed to construct three series of structurally novel and complex steroidal spirooxindoles. All compounds were obtained with high chemoselectivity, regioselectivity, and diastereoselectivity. Among the synthesized steroidal spirooxindoles, compound Ⅲ-3g displayed the best inhibitory activity against SMMC-7721 cells(IC50 = 0.71 μM), 14-fold more potent than 5-FU. Compound Ⅲ-3n and Ⅲ-9d concentration-dependently arrested cell cycle of MGC-803 at G2/M phase and induced apoptosis. Molecular docking studies showed that compound Ⅲ-9d bond to the same active site with the well-known MDM2 inhibitor Nutlin-2, forming H-bond interactions with surrounding residues Lys94 and Thr101.3. Based on the cascade reactions between the steroidal dicyano alkenes and aromatic aldehydes, we synthesized two series of steroidal dienamides. The steroidal β-lactams were efficiently synthesized from the corresponding steroidal dienamides through the base-promoted Aza-Michael addition/aerobic oxidation sequence. Compounds Ⅳ-4d-e and Ⅳ-4j-k represented excellent antiproliferative activity against MGC-803 cells(IC50 < 0.6 μM). Compounds Ⅳ-10 a and Ⅳ-10 b showed remarkable inhibition against SMMC-7721 and MGC-803 cells with the IC50 values of 0.53 and 0.46 μM, respectively. Compound Ⅳ-10 b induced cell death of MGC-803 and morphological changes in a concentration-dependent manner, accompanied with the decrease of MMP.4. Steroid dimerⅤ-3 described herein showed excellent inhibition against human cancer cell lines of different origins and was also toxic to normal cells(L-O2 and GES-1). CompoundⅤ-3 arrested cell cycle of EC109 cells at G2/M phase and induced apoptosis of EC109 cells in a concentration-/time-dependent manner, accompanied with morphological changes of EC109 cells. CompoundⅤ-3 cleaved MDM2, upregulated p53 expression, promoted the release of cytochrome c, and activated caspase-9/-3, finally leading to the apoptosis of cancer cells.5. A series of antiproliferative steroidal hybrids incorporating the DHEA nucleus and different bioactive scaffolds were designed based on the molecular hybridization strategy. The SARs studies indicated that compounds Ⅵ-12a-h with terminal isatin scaffold could selectively inhibit growth of SH-SY5 Y cells overexpressing LSD1(IC50 < 20 μM), correlating with their inhibition toward LSD1. Of these compounds, compound Ⅵ-12 g exerted the best inhibition against SH-SY5 Y cells(IC50 = 4.06 μM). Docking simulations showed that compound Ⅵ-12 g formed strong interactions with the active site of LSD1, while the steroid nucleus occupied the tubulous hydrophobic cavity well. Compound Ⅵ-12 g arrested cell cycle at G2/M phase and markedly induced apoptosis in a concentration-dependent manner, accompanied with decrease of MMP. Additionally,the non-steroidal isatin/triazole hybrids Ⅵ-16a-h were designed based on the structural simplification strategy. Among these hybrids, compound Ⅵ-16 a had good inhibition against different cancer cell lines, comparable to that of 5-FU. Further mechanistical studies indicated that compound Ⅵ-16 a induced apoptosis at least through the intrinsic and extrinsic apoptotic pathways and LSD1 inactivation. Also, compound Ⅵ-16 a inhibited migration of MGC-803 cells. The first steroid-based LSD1 inactivators were described herein.